The sterol composition of three different populations of Ruditapes decussatus from three localities close to each other, but where different environment conditions prevail, was investigated over a period of 14 months. Sterols of adult clams were isolated by thin layer chromatography and identified by gas chromatography/mass spectrometry. In all samples, the major sterol component was cholesterol (>40% of total sterols); other sterols identified were 24-norcholesta-5,22-dienol, 22-cis-dehydrocholesterol, 22-trans-dehydrocholesterol, brassicasterol, 24-methylene-cholesterol, campesterol, stigmasterol, ß-sitosterol and isofucosterol. At each locality, changes in sterol levels followed specific and different patterns, which remained constant for the period studied and allowed the origin of the clams to be distinguished. Stepwise discriminant analysis, based on the percentage amounts of the sterols at each locality, indicated brassicasterol, 22-cis-dehydrocholesterol, cholesterol, 24-norcholestadienol and ß-sitosterol as discriminant variables that distinguish clam populations. Correct identification of the clam origin was achieved in 100% of the samples, demonstrating that sterols can justifiably be used as molecular biomarkers for determining the origin of this bivalve species.